Abstract Systemic lupus erythematosus (SLE) is a prototypic autoimmune disease characterized by autoantibody production and immune complex mediated organ damage. One of the more profound features of SLE is females having a 9:1 prevalence of disease over men. The cause of this gender difference in SLE is multifactorial, including the sex hormones themselves and their receptors. Estrogen acts primarily via its receptors, estrogen receptor alpha and beta (ER1/ER2). Estrogen can also act, however, through non-receptor mediated mechanisms and, in kind, the ERs mediate physiologic functions independent of estrogen. In the previous funding period, we derived ER1 and ER2 knockout lupus prone MRL/lpr and NZM2410 mice. In both strains, the female ER1 KOs developed significantly less proteinuria and pathologic renal disease and had significantly prolonged survival, despite increased serum levels of autoantibodies. These findings led us to postulate that the primary impact of ER1 deficiency in lupus nephritis was on the response of the kidney to inflammation. We derived ER1 KO and ER2 KO mesangial cells from B6 mice and found that ER1 KO mesangial cells had a marked blunted response to TLR 2, 3 and 7 ligands. ER2 expression had no effect on mesangial cell responses as tested. Based on these findings, we hypothesize that the lack of ER1 is renal protective in female lupus mice by blunting the response of mesangial cells to TLR3/7 induced inflammation. We believe this ER1 protective effect is estrogen independent and mediated via TLR3/7 induced phosphorylation of ER1. To test this hypothesis, we propose the following Specific Aims: 1. Determine in vivo the effects of ER1 deficiency on known mechanisms of lupus pathogenesis using sle1, sle3 and sle1/3 congenic mice and bone marrow transplantation of ER1 KO and ER1 WT mice. 2. Define the in vitro molecular mechanisms underlying TLR/ER1 interactions that impact the inflammatory response in mesangial cells assessing the impact of ER1 on the TLR3/7 activation pathways and TLR3/7 on ER1 expression. 3. Define in vivo and in vitro the mechanisms by which ER1 impacts TLR signaling utilizing mutant ER1 knockin strains that affect specific ER1 functions allowing delineation of specific ER1 functions on the immune response. These studies will provide novel insight into the mechanisms by which ER1 deficiency impacts lupus disease expression and further delineate the interaction between ER1 and TLR induced inflammation that may partially underlie the female predominance in lupus. PUBLIC HEALTH RELEVANCE: Relevance Statement Lupus is a disease with increasing prevalence in the Veteran population. It is primarily a disease of women and minorities, who represent an increasing proportion of patients in VA clinics. Of increasing concern is, that despite recent advances in medical care, mortality rates and progression to renal failure continue to increase in lupus in minorities. This proposal addresses one of the key unanswered questions in lupus, that of the mechanism underlying the 9/1 female predominance in disease. We found that estrogen receptor alpha (ER1) is a key mediator of SLE in female lupus prone mice and that ER1 mediates this effect, at least partially, by modulating the inflammatory response to specific TLR ligands implicated in lupus. Further defining the effect of ER1 on SLE will provide new insight into mechanisms of disease and novel approaches to therapy.